There are many environmental applications for biosensors that can provide real-time analyses. For example, one of the most pressing current needs is monitoring for agents of chemical and bioterrorism. These applications require systems that can rapidly detect small organics including nerve agents, toxic proteins, viruses, spores and whole microbes. A second area of application is monitoring for environmental pollutants. Processing of grab samples through chemical laboratories requires significant time delays in the analyses, preventing the rapid mapping and cleanup of chemical spills. The current state of development of miniaturized, integrated SPR sensor elements has allowed for the development of inexpensive, portable biosensor systems capable of the simultaneous analysis of multiple analytes. Most of the detection protocols make use of antibodies immobilized on the sensor surface. For example, the Spreeta 2000 SPR biosensor elements manufactured by Texas Instruments Corporation provide multiple channels for each sensor element in the system. (See, e.g., documents cited in our Information Disclosure Statement and U.S. Pat. No. 5,912,456, Jun. 15, 1997, “Integrally Formed Surface Plasmon Resonance Sensor”.)
SPR biosensors are based on the fundamental Kretschmann design where the intensity of transverse magnetic (TM) polarized light reflected off a thin layer of gold (˜50 nm) on the surface of a prism shows a dependence on the angle of incidence or wavelength of the incident light. See, e.g., Kretschmann E, The determination of the Optical Constants of Metals by Excitation of Surface Plasmons, Z Physik 241:313-324 (1971). A plot of the intensity of reflection against the angle of reflection produces an SPR curve or profile. A minimum in the SPR curve is observed when the frequency and momentum of the incident light are matched with that of the surface plasmons, at which point the energy is absorbed by the surface plasmons (and not reflected). The angle or wavelength at which the minimum of reflection occurs is dependent on the refractive index (RI) of the medium in contact with the outside of the gold layer. Attachment of specific recognition elements on the gold surface (usually antibodies) and passivation of the gold surface to non-specific binding provides a condition for monitoring for the presence of specific targets in real-time. Since the refractive index of protein (RI ˜1.45) is greater than that of usual aqueous buffers (RI ˜1.334), when an analyte of refractive index greater than that of water/buffer and of sufficient size is bound at the surface, the refractive index change is sufficient to result in a change in the position of the minimum of the SPR curve. Instrumentation software can convert the change in SPR minima into refractive index as a function of time, thus allowing the binding event to be analyzed in real time. See, e.g., Davies J. (ed), Surface analytical techniques for probing biomaterial processes, CRC Press, NY (1996).
The first commercial instruments available were both large and expensive and thus not suitable for applications that required portability. Recent advances in miniaturization of SPR technology have made possible the development of portable systems that are adaptable to many different uses. See, Naimushin, et al., Detection of Staphlococcus aureus enterotoxin B in femtomolar amounts by a miniature integrated two-channel SPR sensor, Biosens Bioelectron 17:573-584 (2002); and Naimushin, et al., A portable surface plasmon resonance (SPR) sensor system with temperature regulation, Sens Actuators B 96:253-260 (2003). Such uses include but are not limited to: 1) continuous monitoring of water systems; 2) continuous monitoring of air supply systems, when coupled with a sample collection device that transfers analytes into the aqueous phase; 3) rapid identification of possible agents of chemical or biological warfare; 4) general laboratory analyses of intermolecular interactions, e.g., protein/protein interactions, protein/ligand interactions, and protein/nucleic acid interactions; 4) environmental monitoring; 5) drug discovery; 6) in-office diagnostics; 7) in-emergency-vehicle analyses, e.g., rapid detection of plasma levels of cardiac enzyme levels, and 8) automating protein purification protocols.